光暴露与人晶状体上皮细胞氧化损伤和凋亡的关系及其机制  

Association of light exposure with oxidative damage and apoptosis of human lens epithelial cells

作　　者：魏树瑾[1] 南莉[1] 汤欣[1] 丁娜[2] 黄伟[2] 孙二琳[2] 

机构地区：[1]天津医科大学眼科临床学院天津市眼科医院天津市眼科研究所天津市眼科学与视觉科学重点实验室,300020 [2]天津市泌尿外科研究所,300211

出　　处：《中华实验眼科杂志》2015年第4期300-305,共6页Chinese Journal Of Experimental Ophthalmology

基　　金：国家自然科学基金项目（81270984、61178081）;天津市应用基础与前沿技术研究计划（重点项目）（14JCZDJC36300）;天津市中医药管理局中医中西医结合科研专项课题项目（13134）

摘　　要：背景 研究表明,光暴露可导致晶状体上皮细胞（LECs）的损伤,与活性氧簇（ROS）的累积和自由基反应有关,其发病机制一直是研究的热点.目的 研究LECs光损伤的病理机制.方法 将人LECs系HLEB-3细胞株进行复苏和培养并分为4个组.将20 000 lx光照度的发光二极管（LED）光源置于密封的培养箱并光照培养的细胞6、12、24 h,未行光照射的细胞作为正常对照组.采用流式细胞术检测各组LECs内ROS的表达及细胞凋亡率,分别使用real-time PCR和Western blot技术检测各组细胞中bax和转化生长因子-β2（TGF-β2） mRNA及其蛋白的相对表达水平;采用MTT法检测并比较正常对照组与光照24 h组细胞的增生值（A570）.结果 正常对照组培养的细胞排列紧密,光照后随着时间延长,细胞肿胀,细胞间隙增宽.LECs内ROS平均荧光强度（MFI）、早期凋亡率、总凋亡率随光照时间的延长逐渐上升,各组间总体比较差异均有统计学意义（F=8.276,P=0.036;F=27.878,P=0.006;F=9.333,P=0.028）.光照6h组、12h组和24 h组,bax mRNA的相对表达量较正常对照组分别升高（1.45±0.33）、（5.45±2.58）和（15.86±3.93）倍,TGF-β2mRNA的表达分别升高（1.22±0.12）、（1.79±0.38）和（4.01±0.93）倍,4个组间总体比较差异均有统计学意义（F=13.679,P=0.023;F=13.320,P=0.024）.光照24 h组bax和TGF-β2蛋白的表达量分别较正常对照组升高约1.85倍和2倍.正常对照组细胞增生值（A570）为0.959±0.058,明显高于光照24 h组的0.897±0.093,组间差异有统计学意义（t=-3.130,P=0.003）.结论 20 000 lx的光照可导致体外培养人LECs的氧化损伤和凋亡,bax和TGF-β2可能参与光诱导人LECs的损伤过程.Background Studies showed that light exposure induce the damage of lens epithelial cells （LECs）,and this procedure is associated with the accumulation of reactive oxidative species （ROS） and apoptosis of LECs.However,the mechanism of light damage of LECs is below understood.Objective This study was to investigate the pathogenesis mechanism of light-induced LECs damage.Methods HLEB-3 strains were resuscitated and cultured and then divided into four groups.A light-emitting diode （LED） light source with the illuminance of 20 000 lx was set in a sealed culture box to irradiated the LECs for 6,12 and 24 hours,and the cells were regularly cultured without light exposure in the normal control group.The mean fluorescence intensity （MFI） of ROS and apoptosis of the cells in different groups were tested by flow cytometry （FCM）.The relative expressions of mRNA and protein of bax and transforming growth factor-β2（TGF-β2）in the cells of different groups were assayed by real-time PCR and Western blot,respectively.The growth and proliferation were assessed using MTT as absorbance （A570） and compared between the normal control group and the light-exposure 24-hour group.Results The cells grew well with the tight arrangement in the normal control group,but cellular swelling and intercellular space widening were seen in the light-exposure 24-hour group.The MFI of ROS in the normal control group,light-exposure 6-hour group,light-exposure 12-hour group and light-exposure 24-hour group was 2.85 ± 0.14,3.33 ± 0.21,5.82 ± 1.69,15.83 ±5.81,respectively,showing a significant increase in all the light-exposure groups （F =8.276,P =0.036）.The early apoptosis rate was （0.17±0.13）％,（0.35±0.03）％,（0.74±0.27）％,（3.12±0.47）％,and the total apoptosis rate was （0.27±0.16）％,（0.58±0.14）％,（1.54±0.79）％,（5.22±1.63）％ in the normal control group,light-exposure 6-hour group,light-exposure 12-hour group and light-exposure 24-hour group,respectively,with the significan

关 键 词：晶状体上皮细胞 光损伤 细胞凋亡 活性氧 转化生长因子-Β2 

分 类 号：R329.27[医药卫生—人体解剖和组织胚胎学]